JPS59141549A - Cyclic peptide having somatostatin activity - Google Patents

Abstract

The invention relates to cyclic hexapeptides of the general formula III <IMAGE> in which X represents the radical of an L-aminoacid of the general formula IIIa <IMAGE> (IIIa) in which A and B are identical or different and denote alkyl having 1 to 3 carbon atoms, or A and B together represent a saturated, unsaturated or aromatic monocyclic or bicyclic structure having 3 to 6 carbon atoms, n denotes 0 or 1, and Y represents an aliphatic or aromatic L-aminoacid the side chain of which can be hydroxylated, and their salts with physiologically tolerated acids, and to a process for their preparation and their use and their intermediates.

Description

DETAILED DESCRIPTION OF THE INVENTION Somatostatin (8omatostatin, ) has the formula %
It is a vezutide consisting of 14 amino acids with the formula %( ().

This also occurs in the hypothalamus [5science J Vol.
d, 5upp1. J Vol. 473, p. 15 (1979)], it was also used for the gastrointestinal tract. Somatostatin regulates blood sugar levels, e.g. by suppressing insulin or glucagon, and inhibits growth hormone, TEI.
H, ACTH, prolactin, pancreozymin, secretin, motilin, ■IP1U
Suppresses IP and also suppresses gastric acid secretion by gastrin [: rAn, J, Med, J Vol. 70, No. 61
9-626 (1981)]. These sex tabs (may be used by men as therapeutic agents. Also this 11
It is used to treat disorders of blood sugar levels (eg, diabetes mellitus) because it inhibits the secretion of 11-neurin and glucagon. For example, elevated plasma GH levels that cause acromegaly or psoriasis can be lowered by matostatin. It lowers gastric acid through its gas- and phosphorus-suppressing action and cures pathologies caused by excess gastric acid, such as gastrointestinal bleeding. For example, Verne
r)-Morrison syndrome (ViP-producing tumors), or Zollinger
)-Ellison (gl]-180n) syndrome (Gust IJ
The growth of lumone-producing lung tumors, which are the basis of lumone-producing tumors), can be inhibited by somatostatin.

However, somatostatin is very easily administered and therefore it only makes sense to administer it as an injection. The search for long-acting somatostatin analogs is warranted. By replacing Trp with D-Trp, a somatostatin analog with even more potent action was obtained, which inhibits the secretion of growth hormone and insulin by somatostatin. approximately 8 times and approximately 6 times more potently inhibits glucagon (rEiochem
, BiophyB, ReS, Commun, J Vol. 65, pp. 746-51 (1975)]. D-Tr
Formula ■P in which PhO is replaced by Pro in addition to p
ro-Phe-D-Trp-Lys-Thr-PhO (
When shortened to a cyclic hexabetide with II), it already shows a strong and sustained nomatostatin action [r
Nature J Vol. 292, pp. 55-58 (198
1 year)].

It has now been found that the action of matostatin can be further enhanced by substituting turoline of formula (2) with a more lipophilic heterocycle.

The present invention relates to the general formula III X-Phe-D-Trp-Lys-Y-Phe
(Ill) [In the formula, represents a saturated, unsaturated or aromatic monomial or bicyclic structure having 6 carbon atoms;
and n represents LO sweet or 1], and Y represents an aliphatic or aromatic L-amino acid whose side chain may be hydroxylated. Hexa relates to otide and its salts with physiologically acceptable acids.

The present invention further provides the general formula (i) a-X-a-Phe-a-D-Trp-a-Lys(R
”)-a-Y-a-Phe (IV,) in which X and Y have the meanings defined above, R1 represents a protecting group for the ε-amino function and 5 representing a chemical bond and one representing 10H0H-) is cyclized by known peptide synthesis methods and then the protecting groups present are removed by suitable methods. It also relates to a process for producing a compound having formula (1), characterized in that it is removed by:

It is understood that the peptide of formula (1) above specifically includes compounds of formulas IVa to lVf below.

H-X-Phe -D-Trp -Lys (R1 piece Y
-Phe-CIH(IVa)H-Phe-X-Phe-
D-Trp-Lys(R,1)-Y-OH(■b)H-
Y-Phe-X-Phe-D-Trp-Lys(R' fragment OH(IVc)H-Lys(R')-Y-Phe-X
-Phe-D-Trp-OH(■d)H-D-E'rp
-1i, yg (R' piece Y-Phe-X-Pbe-OR
(IVe)H-Phe-D-Trp-Lys(R,'
, )-Y-Phe-X-0) ((IVf) in the above formula, B
1, X and Y each have the meanings defined above.

The present invention is based on the shortened formula (in the formula, a, Rj, X and Y are I)
'J ri[',, has the meaning and in the formula five of the groups a are chemical bonds and one more i: -O
R+Z-, in which R stands for alkyl having 1 to 6 carbon atoms, preferably methyl), and is characterized in that the ester is alkaline saponified and then the two groups are removed by hydrogenation. The present invention also relates to linear hexabeotides having the formula (1) in which a, X, Y and B1 have the meanings given above, and methods for their production. In detail, the esters of (2) and (2) are understood to be compounds of the following formulas V a - V f.

Z -X-Phe -D-Trp-Lys (R1 piece Y
-Pb-e-OR(Va)Z-Phe-X-Phe-D
-Trp-Lys(R1)-Y-OR(Vb)Z-Y-
Phe-X-Phe-D-Trp-Lye (R1 piece OR
(Vc)Z-Lys(R1)-Y-Phe-X-Phe
-D-Trp-OR(Va)Z-D7Trp-Lys(
R')-Y-Phe-X-Phe-OR(Ve)Z
-Phe-D-Trp-Lys (R1 LeY-Phe-
X-OR(Vf) where X, Y, R and R1
has the meaning given above. R1 preferably represents 13oc.

Synthesis of compounds of formula Va-Vf was carried out by Norifield (Me
rrifield) or by the classical route in solution. Standard methods are e.g.
Mr. roes The Peptides Analysis
sis.

5ynthesie, Biology J Volume 1 rMa
jor Methods of Peptide Bon
d Formation, Part A J (19
1979).

If cyclohexa is a notide, there are six possibilities for cyclization of various chain peptides into the same cyclopeptide. Since tritophane tends to form by-products during the removal of protecting groups with acids (e.g. removal of the Boc group) in solid-state methods, in such cases tritophane can be added to the final amino 112
It is convenient to add it as Using the solid-state method, for example, the general formula ■ HD-Trp-Lys (Z)-Y-Phe-X-P
Hekya azide having the formula he-0-R2 (■) (where X and Y have the above-mentioned meanings (- and R2 represents a solid resin) is prepared. As a protecting group for the ε-amino function of lysine, A urethane film-retaining group (7 or 0 benzylmexycarbonyl group) is used. The β-hydroxy group of threonine can remain free.

Hexapeptides attached to the resin via ester functions are removed using hydrazine. A modified hydrazide is formed, which is preferably cyclized after conversion to an azide. The isolated cyclic peptide is purified by total chromatography. The diastereoisomers present are also separated. Removal of the protecting group from the benzylic form by catalytic hydrogenation provides the cycloazido according to the invention.

In classical R azide synthesis, for example, the 2-group that can be removed by catalytic hydrogenation or the secondary amine Kj can be removed by catalytic hydrogenation.
~Fluorenylmethyloxycarbonyl group (Fmoc)
is used as the amine protecting group, while the ε-amine group of lysine is preferably protected by a BOC group. General formula V
The a-Vf azide is synthesized stepwise.

Alka IJ ff of ester (good-IL< is OMe)
oxidation produces free acid. The α-amine protecting group is then removed and the peptide, with its N- and C-termini free, is cyclized by peptide chemistry methods. The protected ring azide is purified by chromatography. The tertiary butyl group is conveniently removed using trifluoroacetic acid with addition of 1,2-dinotylcethane.

The racemic amino acid having the formula 111b (wherein A, B and n have the meanings given above) is, for example,
P-'A) No. 50800, No. 31741, No. 5
No. 1020, No. 49658, No. 49605, No. 29488, No. 46953, and No. 5287
It is known from the description in the book No. 0 Mei'IftII. Tetrahydroisoquinoline-6-carboxylic acid is j J, Am
Er.

Chem, Soc, J No. 7 [Volume 1 No. 182 Negative (19
(1948). decahydroisoquinoline
3-carboxylic acid from European patent application CFP-A) no. 52870 and 2,6-sihydro[1H]
-Indole-2-carboxylic acid in U.S. Patent No. 4.303
, 583. Sis.

Echino-octahydro-[1H]-indole-2-carboxylic acid, cis, exo-octahydro-2-cyclo(b) virol-2-carboxylic acid and cis, exo-azabicycloC5,3,0) decane- 3-Carboxylic acids are inter alia German Patent Application No. P 315169
This is the objective of No. 0.4. German patent application no. P 322
No. 6768.1 relates to, inter alia, cis, endo-octahydro-cyclopenta[b]virol-2-carboxylic acid, and No. P3246503.3 relates to, inter alia, cis, endo-azabicyclo[5,3,0]decane-6-carboxylic acid. Regarding P 3210496. No. O is above all Sis.

Endo- and cis-, exo-2,3,3a, 4,5.
7a -hexahydro-[IF(]]indole2-
Regarding calfone, also No. P 3300774.8 relates inter alia to diethylturorin and also No. P
No. 3242151.6 relates inter alia to 6-asadolycycloC5,2,1,02.6]decane-4-carboxylic acid. Cis, endo-octahydro-[1H]indole-2'-L-carboxylic acid is known from European Patent Application (BP-A) No. 37231; amino acid Y
For example, L-alanine, L-serine, L-threonine,
It can be L-valine, L-leucine, L-(soloucine, L-phenylalanine or L-tyrosine.

As Bo Chiang Ki P1, 1P was created by Mr. Bodanszky et al.
eptideSynthesis J 2nd edition (1976
The conventional JHI honors group described in 2010) may be used. Alkanoyl having 1 to 6 carbon atoms, 6-butoxycarbonyl and benzyloxycarbonyl are preferred.

The invention further relates to the use of the compounds of Part 2 as medicaments, pharmaceutical formulations containing these compounds, processes for their preparation and their use as medicaments.

Belotide according to the invention has somatostatin inhibitory properties, but works much longer in lower doses. For example, peptides reduce gastric acid by an ED5o of approximately 5o7/9 after intravenous administration to rats. Decrease in stomach acid is 2
Persistent over time. In comparison, somatostatin has no effect on gastric acid at an intravenous dose of 3 μf/Kf.

Only infusion of somatostatin brings about a decrease in gastric acidity. The ED5Q in growth hormone suppression is also 4I at about 2-1 oμf/intravenously. In a study on the synergistic effect on the hypoglycemic effect of insulin in adrenally imprinted rats, for example, cisr+ -(AOc-Pl
]e-,D-Trp-LyEl-'7al-J)he
) is cyclo-CPro-]'be-I)-Trp-Ly
E- showed at least twice as strong an effect compared to +>-Thr-t'hθ). Aoc analogs have a cutoff of 0. Q7/
Even a dose of 7S'/V, g lowers blood sugar levels by 24% +
0.318 per minute, whereas]'rO analogues
Even an amount of 1+ri/I'y had no effect at all.

The novel compounds of the present invention can also be applied orally and intracranially. However, in that case, the absorption capacity is poor and a considerably large amount is required.

Due to its co-activation with somatosfutin, the novel compound 1d: somatostatin can be used in all cases where injections show a favorable effect. For example, gastrointestinal bleeding, gastric ulcers, and the production of hormones that can be suppressed by statins.
], 1 day on) syndrome, Ferner-Morin syndrome or fc is used in the treatment of tumors such as insulin- or glucagon-producing tumors, hormone-dependent tumors when the corresponding hormone can be suppressed by somatostatin, certain leukemias, e.g. Metabolic disorders with elevated hormone levels that can be suppressed by somatostatin, such as rheumatoid arthritis, when plasma insulin and plasma growth hormone are too high, acromegaly, dry nails, diabetes (glucagon suppression), chondrosarcoma and shock state.

The effective dose in humans is 0.1 to 10 μW/f when administered parenterally.
And about 1 to 100μ when used intranasally! 7/Kf.

Since somatostatin does not suppress externally imposed insulin, combination preparations with insulin are preferred in the case of diabetes.

Example 1 General formula Boa-D-Trp-Lye(Z) -Y-P
Linear carrier bond B with he-X-Phe-0-R
OC-hekiza preparation of tutide hydroxymethylated Merrifield
field) Resin 24f and approximately 360 ml of CH2CL
2 and into a stirred suspension 13oc-
Phe-oH15,9y, dicyclohexylcarbodiimide 12.4F and 4-dimethylaminopyridine 7,
4 Add Y. Incubate overnight to ensure as quantitative a reaction as possible. The solid residue is filtered and washed (CH2Cl250 m/X I CH2C42/methanol (1°1) 50 me x 4, CH2Cl250 t
nlX2).

Next, the unrestored OH function of the resin is CH,,C4C4
The image is maintained using 3 ml of benzoyl chloride in 2200 and 2.4 mg of pyridine as base. Bo
Solid phase synthesis shown in the following steps is carried out using each of these resins 41 loaded with a-Ph and e.

Time Quantity Number of steps (min)
1:1) 3 2 4 40 CH2Cl2/methanol (1:1) 4 3 5 50 CH2Cl25
10 in 3 5 50 cH2cz2
% diimpropylethylamine 6 5 3 50 cH2c
z27 1 240 60 CH2C
42, IQmMol Boc-amino acids, 10 mMo1 dicyclohexylcarbodiimide, 10 mMo1 1-hydroxybenztriazole 8 4 5 50 CH2Ct2/M
eOH (1:1) 9 2 3 50 CH2Cl2 Step 70 hours The stated time was greatly exceeded. This is because most of the time the equipment must be turned on at night.

Check for completion of the coupling was performed after step 9 using picric acid or chloranil in toluene. After the final wash in step 9, the loaded Norifield resin was completely vacuum dried.

Yield 5.9-6.2t.

Example 2 General formula Boa-D-Trp-Lys(Z)-Y-Ph
Boc-hexa with e-X-Phe-NH-14H2 Preparation of Wotide hydrazide Boc-hekiza with e-X-Phe-NH-14H2 Wotide 67 coupled with a carrier was dissolved in DMF 100+++/
5 ml of anhydrous hydrazine hydrate are added and stirred at room temperature for 2 E:I 1) Jl. The residue is filtered with suction, washed thoroughly with dimethylformamide and methanol and the p-liquid is concentrated to dryness. To completely remove excess hydrazine, the residue is dissolved several times with methanol/toluene (1:1) and removed again on a rotary evaporator. The residue is soaked several times with a small amount of water to remove the benzoyl hydrazide, finally filtered and dried over P2O5 under an oil pump vacuum. The crude yield varies from 1.0 to 2.21, depending on the type of peptide.

The crude R peptide was chromatographed on silica gel [solvent: CH2Ct2/MeOVHOAc (100:
8:5)].

Example 3 General formula HD-Trp-LyF3(Z)-Y-Phe
Preparation of Boc-hexabeptide hydrazide with -X-Phe-NH-NH2 Substantially dissolve 1 mmol of Boc-hexabeptide hydrazide in about 2 ml of methanol and add 0-4 N ICtl in methanol. .

The mixture is stirred for about 15 minutes to exclude moisture, 2 and then concentrated to dryness on a rotary evaporator. The hekylyhydride thus obtained is immediately used in the next step without further characterization of othydrazidobis-hydrochloride.

Example 4 General formula 'X-Ph, e-1)-Trp'-Lye (
Cyclization of hexapetide hydrazide dihydrochloride to a compound with Z)-Y-Phe in DM
Dissolve in 25 ml of F. To the stirring solution at -15° C., 4 N HCto, 67° C. in dioxane is added and then 0.18 td amyl nitrite. Leave at low temperature for about 40 minutes 1. and then preheat the reaction mixture to -20°C.
The cooled DM, F,2 is transferred into 1- and neutralized with 0.81 ml of diisoturobyl ethylamine.

The whole mixture was left at about 0 °C for 2 days, then brought to room temperature and stirred for a further 6 days. The residue remaining after removing the solvent was dissolved in methanol/water (1:1). and stirred for 24 hours with approximately 8007 nl of mixed bed ion exchanger. After stripping off the ion exchanger and concentrating the aqueous methanol phase, an amorphous residue remains which is subjected to semi-preparative HPLC on silica gel [eluent: CH2Ct2 /H2C-2
/acetic acid (100:5:0.5)).

Example 5 Preparation of Clopeptide The main fraction obtained by HPLC to remove the ε-2=protecting group of lysine is dissolved in methanol and hydrogenated for about 2 hours in the presence of a palladium catalyst.

After stripping off the catalyst, it is concentrated under vacuum and chromatographed again on silica gel (methylene chloride/methanol/acetic acid/water (70:30:1.5:6)).

Table 1 Physical data of Boa-hexaR otide hydrazide'J
,”1cThr 2.00 1.03 1.00 0
．． 97 7.3゜01cThr 2.00 0.9
4 0.92 1.03 -1'5.8゜AocThr
1.99 0,97 1.00 1.04 -1
1.1゜ Characteristic NMR data (δ value) of the cyclic compounds shown in Table 2 Tie
Thr 10,72 8.60 7.54. 5.1
B 5.0 1.0B8.59 8.68 7.80 01, c Thr ILl, 130 8.78 7.
63 5, OO, 9552 8゜08 Aoc Thr 10.70 8.5B 7.47
5.0 1.028.68 8.30 7.79 General formula X-Ph, e-D-Trp-Lys-Y-Ph
The final product with e is benzyloxycarbonyl (1
) shows the same characteristic NMR data except that there is no signal for the group. The abbreviations in the table are as follows.

Tic = tetrahydroisoquinoline-6-carboxylic acid 01c = cis, endo-octahydroindole-2
-Carboxylic acid Aoc-cis-octahydro-cyclo[b]-pyrrole-2-endo-carboxylic acid example 6 cisO-(Aoc-Phe-D-Trp-Lys-Va
l-Phe)a) Z-Phe-Aoc-OBzl Salt of Z-Phe-OH and H-AO(!-QBZI 1a5
1 is dissolved in dimethylformamide 100. To this, 0℃TeHOtBt 4.831i' and DCC7
, Of is added.

Stir at 0° C. for 1 hour and leave at room temperature overnight.

The next day, the precipitate is filtered with suction and the P-sin is concentrated.

Remaining W? R things! "Dissolved in acid ester and water, saturated NaHCO3 solution, I N r" 2s○4, saturated Na
Mix with T4CO5 bath solution and water. The acetate bath is dried over Na2804 and concentrated. The yield of oil was 1619°. Purification was carried out by chromatography on silica gel using notylene chlorite/tutanol (9,7: 0.3 v/v). Yield of clear oil 1541i'
, DC [methylene chloride/methanol (9,7:0
．． 3).

h) Z-Phe-Aoc-OH dioxa/water (822) Z-P in 60 ml
2 in a solution of he -Aoc-QBz]j 5.4 f
Add 14.5 ml of N NaOH. Let stand at room temperature for about 20 hours, evaporate with 1N H2SO4 and concentrate. Add the residue to water (pour the solution into 1N H2
Acidify to T)H2~B with SO4. The precipitated oil was extracted with acetate, and the acetate bath solution 'f: Na2
Dry with SO4 and concentrate. Foam yield 13.7
g.

c) Z-Phe-Aoc-OI in Z-Phe-Aoc-Phe-OMe dimethylformamide 50-
13,71i', H-Phe-OMO・HCl6.77
r and HOBt4.46fi! N in a solution of KO at °C
- ethylmorpholine 4.02 ml and]) CC6,
Add 9V. Stir at 0°C for 1 hour, then leave at room temperature overnight. Work up as in Example 6a. Foam yield 1.8.85'.

Purification as in Example 68 yields a transparent oil 16, Or
get.

a) H-Phe-Aoc-]-'he-OMe・H
CtZ-Phe-Aoc-Phe-OMe 16. O
ffDissolve in methanol. To this, 10% Pd-
C catalyst is added and hydrogenated in an automatic titrator at pH 4.5. After the hydrogenation is completed, the catalyst is suction filtered through diatomaceous earth, concentrated, and the residue is triturated with diethyl ether. The precipitate is filtered off with suction and dried.

Yield - It, 8.67, melting point 79-91°C, [α],
, =-1-26,9' (c?1, methanol 7).

e) Z-Va1 he-AoC-Phe-OMe dinotylformamide 6-Phe-OMO-HCl2. O Y, Z-Val-O
H2. [J 1 6! / and [10131; i.
N-ethylmorpholine 1 at 0°C in a solution of 1 4 9
03rn7! and add 1.76 fl of DCC.

Leave at 0°C for 1 hour, then at room temperature overnight. Example 68
Work up and purify in the same manner as above. Oil yield N4.
2ri.

i”) H-Val-Phe-Aoc-Phe-
OMe-HCtZ-Val-T-'he-Aoc-Ph
e-OMe 4.9 5 9 in methanol r4I
Q'F and catalytic hydrogenation as in section d. Yield 6.619 (oily substance).

ry,) Z-D-Trp-Lye(BocJ-OH
H-Lys (
Boc)-OH 1 1.8 9
OBt 6. 8 7ri and Z-IJ-Trp-OT
Add 5 g of cp. - Stir overnight 4b1 and concentrate. The residue is triturated three times with petroleum ether and countercurrently partitioned in three stages between acetic acid ester and saturated IJ Na HC 0 5. Combine the acetate ester phases and KH
Shake with SO4/2SO4 solution, dry with Na2SO4 and concentrate. Foam yield 4912.

Further purification is achieved by chromatography on silica gel 4001. First elute with methylene chloride and transfer the material to methylene chloride/methanol/water [16
:4:0.3(v/V)].

Concentrate the bath liquid and dry under a trough. Amorphous foam yield 31.1 f.

h) Z-D-Trp-Lye(Boc)-Val-
H-Val-Phe-Aoc-Pk+ in 20 ml of Phe-Aoc-Phe-OMe dimethylformamide
e-OMe・HCl 3.31, Z-D-Trp-L
yaCBoc)-OF+ 5.12! i' and Wo
oJ:it Elf, 1 in o'c in solution of 9 f
Liech/L = Morpholine 0.7 ml and JJC
Add C1,14V. Work up as in Example 6a.

Yield f: (4,611i', melting point 117-119°C, [α
, I1. =-24.2° (C-1, methanol).

t) Z-1,3-Trp-Lye(Boc
)-Val-Pbe-Aoc-Phe-0cho■Z-D-
Trp-Lys(Boc)-Val-Phe-Aoc
-Phe-OMe 2.67 to dioxane/water (8:2
)Mixture 2 S Make a lewd child inside me. I to this
N Na, OH 57 and at room temperature 117. '?
Leave it for a while. Then concentrate the solution using 12SO4 medium. The residue is Example 6b
Post-process in the same way. Yield: 2.22. Decomposed at 118°C or higher, [α] -25.9° (c; 1, methanol).

k) +4-D-Trp-Lys(Boc)-V
a,]-]Phe-Aoc-Ph,e-OHz]stopTr
p-Lye(Boc)-Val-Phe-Ao, c-P
he-OR2,22 was dissolved in 90% acetic acid and P
After adding the d-C catalyst, hydrogenation is carried out. After the hydrogenation is completed, the catalyst is filtered through a clarification p-filter, and the p-liquid is concentrated. Grind the residue with water. This suspension (pH 3,1)
Add NaHCO3 solution (about 2.5 me in total) under stirring until the pH is about 7. The precipitate is filtered off with suction and washed thoroughly with water. Yield 1.12 f, [α]. =-43,
5° (c-1, methanol), decomposes above 168C.

1-) Schiff o -(Aoc-Phe-D-Trl:
+-Lye(Boc)-val-Phe)HD-Tr, p-Lye(Boc)-val-Phe) in 200 ml of dimethylformamide
Boc )-Val-Phe-Aoc-Phe-OH4
Niethylmethylphosphinic anhydride (50%) in a solution of 81.8 mg 0.25 m7! and (slowly add a solution of 0.4-N-ethylmorpholine in dinotylformamide 10- with vigorous stirring. After about 2 hours the solution is concentrated and the residue is triturated with water. Yield 0.57°. At 51°C, perform chromatography on a gel using notylene chloride/methanol/water (1800:280:20). Collection f11'f 200"2
゜m) Cyclo-(Δoc-Phe-D-Trp-Ly
e-Val-Phe)cyclo-(Aoc-Phe-D
-Trp-Lys(Boc)-Val-Rle)20
0mLj to trifluoroacetic acid/water/1,2-ethanedithiol (4,5me: 0.5m/!: 0.5ml
) is dissolved in a mixture consisting of: Leave at room temperature for 90 minutes, concentrate and partition the residue between water and methyl 6-butyl ether. The aqueous phase is adjusted to pH 3.6 using a weakly basic ion exchanger (acetate type) and freeze-dried. Yield 126.7#+go amino acid analysis (6N
Hydrolysis for 24 hours at 120°C in HC1): Val (
0,97), Phe (2,05), Lys (1,0
0), Aoc (0,95B” peptide base content 281%
).

1'rp is destroyed under this condition. This R notid U
Vs and trP showed absorption at 277 nm, which is characteristic of Trp.

Example 7 Schiff c+ -(D-Trp-hys-Thr-Ph, e
-Aoc-Phe) a) 13oc-Aoc-Phe-
OBzlBoa-Aoc-OH9,8t, HCt-H
-Phe-OBzl 11.2 f.

HOBt 5.2 f and toI-ethylmorpholif
Dissolve 4.9 ml in 100 ml DMF and
C7,9ii' is added (7) and stirred at room temperature for 14 hours. The precipitated DC-urea is sucked off, the p-liquid is evaporated under vacuum, the residue is taken up in 200 m7 of acetic acid ester and citric acid solution is added. and extracted with NaHCO3 solution.

Concentration of the organic phase leaves an oily residue. Yield: 20 liters.

b) H-Aoc-J'be-QBzlTFAcO
20 g of the Boc compound obtained in Example 7a are dissolved in 50 ml of trifluoroacetic acid. After 45 minutes it is concentrated under vacuum. An oil is again obtained. Yield 22 ii'.

c) Boa-Phe-Aoc-Phe-OBzI
T-1-Aoc-Phe-OBz14FAcOH22W
, 11.5 g of Boc-Phe-OH, 5.6 ml of 1-lythylmorpholine and 5.9 Y of HOi3t are dissolved in 150 ml of acetic ester. DCC9,l
After adding Ff, the mixture is allowed to react at room temperature for 18 hours. It is filtered with suction, the organic phase is washed with citric acid and l'JaHcJ solution, dried over solid sodium sulfate, evaporated and concentrated. Yield i22.4.

6,) H-Phe-Aoc-Phe-OBl, 4
FAcOB Example 7 c “”C? ? ) Boc compound 22.4? 150ml of trifluoro vinegar. dissolve in it. After 1 hour, concentrate at room temperature. Yield: approximately 262゜e) Z-Lys(Boc)-Thr(tHu)-
Phe-Aoc-Phe-OBzllll-Phe-Ao
c-Ph, e-OBzl・TFAcOH, 5,3f,
Z-Lys(Boc)-Thr(tBu)-OH2,7
8f/,) (0.7 ml of OBt O,7y and N-ethylmorpholine are dissolved in 50 mA of acetic acid ester. After adding 0.7 g of DCCl, react overnight at room temperature. Using citric acid and bicarbonate solution. After shaking, drying and evaporation. The remaining brown oil is filtered over 400f of silica gel (s:to2-60, Merck & Co. product) using the solvent system C'HCHV/MeOH (13:1). Yield 5 .1f0f) H-Lys(Boc)
-Thr(tBu)-Phe-Aoc-Phe-OH2
-Compound (product of Example 7e) 57 in methanol 150-
After the addition of 0.42 Pa/C (5% Pd), hydrogenation is carried out. After absorption of hydrogen has ended, it is filtered and concentrated under vacuum.

Silica gel column solvent system CHC63/MeOH/HA
Chromatograph using cO (50:20:5). Two types of compounds were obtained with a yield of 2.17°, and the compound with a higher Rf value was used for further reaction.

g) Z-D-Trp-Lys(Boa)-Thr(
tBu)-Phe-Aoc-PheOH 11-Lye(&)c)-Thr(tBu)-P
he-Aoc-Ph θ-OH300mg, Z-1)-
Trp-OTcp 190 mg, HUBt 50 m'
i), 50 pt of N-ethylmorpholine is dissolved in 10 mf4 of acetic acid ester and left at room temperature for 48 hours.

This is evaporated (7) and chromatographed on silica gel 15o7 [solvent system: CHCt3/MeCIH (
5:1)]. Yield 260■.

h) H-D-Trp-Lye(Boc)-Th
r(tBu)-Phe-OH2 derivative (example 7 g of product) 260 m''/ is dissolved in MeOI425-,
Add Pd/C100 and hydrogenate. The catalyst is removed and concentrated under vacuum. Yield: Approximately 240 pieces.

j) Schif o -(D -Trp-11+ys(Boc
)-Thr(tBu)-Phe-Aoc-Phe
) Linear Belotide (Product of Example 7h) 700 nry t
]]+11F' 350 pt ethylnotylphosphinic anhydride and 0.1 ml N-ethylmorpholine dissolved in 25 ml and under stirring! Add. Once reacted, evaporate under vacuum and remove the residue from silica gel 5
01 Upper solvent thread CH2CA2/MeOH/H2(J (9
Chromatograph using 0:15:υ. Yield 1500W0j) Cyclo-(D-Trp-Lys-T
hr-Phe-Aoc-Phe) Protected cyclo-hexapeptide Product of Example 71) 500~ is dissolved in trifluoroacetic acid 5-. After 45 minutes it is concentrated under vacuum and precipitated with ether. Take up in a little methanol and precipitate once more with ether.

Yield: 450q. Amino acid analysis: Lye (1,03)
, Phe (2:0), Thr (1,01), Ao
c (0,99), including f1= :B □%.

The synthesis scheme in Example 7 was as follows.

Agent Patent Attorney Yamashita Japan Page 1 Continued 0 Inventor Rolf Geigel Federal Republic of Germany Day-6000 Frankfurt am Main 50 Heinrich Breichelstraße 33 (7 ■ Inventor Jürgen Tart Sandu Germany Federal Republic Day - 6240 Königstein/Taunus am Heideplacken 22

Claims (1)

[Claims] 1) General formula ■ [In the formula, X is the same or different alkyl having 1 to 3 carbon atoms, or and B together represent a saturated, unsaturated or aromatic monocyclic or bicyclic structure having 5 to 6 carbon atoms, and n represents O or 9 to L- a residue of an amino acid and Y represents an aliphatic or aromatic L-amino acid whose side chain may be hydroxylated] and its salts with physiologically acceptable acids. 2) A compound having the formula (2) according to claim 1, wherein Y is Thr. 3) The compound having 2) according to claim 1, wherein X is a residue of tetrahydroisoquinoline-3-L-carboxylic acid. 4) The compound having 2) according to claim 1, wherein X is a residue of cis, endo-octahydro[1H]indole-2-L-carboxylic acid. 5) X is cis, endo-octahydro-cyclo is (
b) A compound having 2) according to claim 1, which is a residue of virol-2-L-carboxylic acid. 6) General formula ■ a-X-a-Phe-a-])-]Trp-a-Lys
R-a-Y-a-P1]e(IV) (wherein X and Y
has the meaning defined in claim 1,
Blt represents a protecting group for the ε-amino function, and five of the groups a in the above formula represent chemical bonds, and one represents a -O
A linear hexa having R + H-) is characterized in that the otide is cyclized by the known otide synthesis method and then the protective groups present are removed by a suitable method. Method of manufacturing compounds. 7) Hexabetide having the general formula (3) in which a, X, Y and R1 have the meanings defined in claim 6 above. 8) A pharmaceutical formulation containing a compound having formula 11 according to any one of claims 1 to 5.